Cloning of Fab Gene of an Anti-Human Bladder Cancer Monoclonal Antibody and Its Expression in E. coli
- VernacularTitle:抗膀胱癌单抗Fab段基因的克隆及表达
- Author:
Lijun ZHOU
;
Yan WANG
;
Yin BAI
;
Hairong ZHANG
;
Lizhang YU
- Publication Type:Journal Article
- Keywords:
anti-human bladder cancer mAb;
antibody variable region gene;
Fab
- From:
Chinese Journal of Cancer Biotherapy
1995;0(02):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective: To clone the Fab gene of a monoclonal antibody (mAb) BDI against human bladder cancer and its expression in E. coli. Methods: Fd and K genes of mAb BDI were cloned by RT-PCR and inserted into an Fab expression vector. Phage displaying Fab and soluble Fab were expressed in E. coli. The N-terminal sequence of VH region was corrected by PCR mediated mutagenesis. The antigen-binding characteristics of the Fab were tested by ELISA and immu-nohistochemistry. Results: Fd and K genes were cloned into the expressing vector p3MH and the phage displaying antibody and soluble Fab were expressed in E. coli, which showed weak binding activity to bladder cancer cells. Correction of the N-terminal sequence of the VHimproved the biding activity dramatically. The feasibility of the application of the Fab in phage antibody library screening was confirmed by a simulated panning procedure. Conclusion: The Fab gene of an anti-human bladder cancer mAb was expressed in E. coli. The importance of the N-terminal sequence on antibody binding activity was suggested.
